The objective of the project is to study the role of heterotrimeric G proteins and their coupled receptors in normal cell growth and oncogenesis. We have genetically engineered NIH 3T3 mouse fibroblasts to express the family of human acetylcholine muscarinic receptors (mAChRs). Using this model, we have shown that genes for mAChRs subtypes coupled to the activation of phosphatidylinositol (PI) hydrolysis can act as ligand-dependent oncogenes, whereas those coupled to the inhibition of the adenylyl cyclase (AC) are not. Furthermore, expression of GTPase deficient alpha subunit of Gq, an activator of PI- phospholipase C, induces focus-formation in NIH 3T3 cells. In contrast, expression of activated Gi2, an inhibitor of AC, fails to transform the same cells. The region of the mAChR that confers transforming potential has been mapped by a receptor chimera approach. We have found that the region responsible for coupling to other signal transduction pathways, such as the activation of phospholipases A2 or D, is also responsible for transforming activity. Furthermore, available evidence suggests that activation of the phospholipase A2-eicosanoid pathway is strictly necessary for mitogenesis as well as transformation induced by receptors coupled to G proteins, but not by activated tyrosine-kinase receptors. Mitogenic signalling through G protein-coupled receptors does not require Ca2+-dependent protein kinase C, but involves rapid tyrosine phosphorylation of cellular proteins, affects the functions of p21ras, the c-raf serine-threonine kinase, and induces expression of certain early-responsive proto-oncogene. Work is in progress to determine the molecular basis for these interactions.